1. Field of the Invention
The present invention pertains generally to high-resolution molecular spectroscopy and imaging devices and methods, and more particularly to an apparatus and method for ex-situ NMR spectroscopy and imaging.
2. Description of the Background Art
Nuclear magnetic resonance spectroscopy is a sensitive tool for studying the physical, chemical and biological properties of matter at a molecular level. One-dimensional and two-dimensional NMR imaging techniques are routinely used by chemists to determine the structure of simple and complicated molecules and such techniques are replacing traditional x-ray crystallography as the preferred method for determining smaller protein structures of 25 kDa or less.
The phenomenon called nuclear magnetic resonance (NMR) occurs when the nuclei of certain atoms are placed in a static magnetic field. The nuclei of the atoms of elements with an odd atomic number possess spin (l) and have a nuclear magnetic moment. As the positively charged nucleus spins, the moving charge creates a magnetic moment.
When no external magnetic field is applied, the magnetic moments of nuclei are aligned randomly. However, if the nuclei are placed in an external homogenous magnetic field (B0), the magnetic moments will either align with the external magnetic field or in opposition to the magnetic field. The alignment of the groups according to one of the two possible orientations follows Boltzmann""s statistics and results in a population imbalance among the different energy levels and a net nuclear magnetization M. Accordingly, there will be slightly more nuclei at the lower energy level than at the higher energy level.
Because nuclei behave like magnets, the nuclei have a lower energy state when aligned with the applied magnetic field than when the nuclei are opposed to the magnetic field. In an applied magnetic field, the axis of rotation will precess around the magnetic field. A nucleus in the low energy state may transition to a high-energy state by the absorption of a photon that has an energy that is exactly equal to the energy difference between the two energy states. The energy of a photon is related to its frequency by Plank""s constant. The frequency of the photon and the equivalent frequency of precession are referred to as the resonance or Larmor frequency.
Thus, it is possible to make magnetic dipoles xe2x80x9cflipxe2x80x9d from the low energy, more stable alignment to the high energy, less stable alignment by supplying the right amount of energy. The energy necessary to make this transition depends on the strength of the external magnetic field used and is usually in the range of energies found in radio waves. Therefore, the nuclei can absorb and reemit energy at characteristic radiofrequencies (rf). Furthermore, energy will be absorbed by the same nuclear species at slightly different frequencies depending on the molecular environment of the nucleus of a particular atom.
The precise resonant frequency of the nuclear species is dependent on the magnetic field at the nucleus that will vary depending on the types of nuclei and the bonds in the molecule involving the nuclei. This characteristic variance in the resonance frequency depending on the chemical environment of the nucleus is called the chemical shift (xcex4) and can be used to deduce the patterns of atomic bonding in the molecule. Chemical shift is the frequency difference between the observed resonance and a resonance from a standard compound and is usually reported in parts per million (ppm) of the mean resonance frequency.
In the typical NMR experiment, the sample is placed in a homogeneous static external magnetic field (B0). By convention, B0 and the net magnetization vector (Mz) reside on the z-axis at equilibrium. Also by convention, a rotating frame of reference rotating around the z-axis at the Larmor frequency allows B0 and net nuclear magnetization M to appear static i.e. the xxe2x80x2 and yxe2x80x2 axes rotate about the z-axis.
Accordingly, the applied radio frequency (rf) pulse has a stationary field vector in the xy plane within this reference frame with a direction governed by the phase of the radio frequency. The application of an rf pulse along the x-axis rotates the nuclear magnetization vector towards the y-axis at an angle that is proportional to the duration and intensity of the rf pulse. A pulse that is of sufficient duration and intensity to rotate the magnetization vector clockwise 90 degrees about the x-axis is termed a 90xc2x0 or xcfx80/2-pulse. Similarly, a 180xc2x0 pulse will rotate the magnetization vector 180 degrees and is called a xcfx80 pulse.
Predictably, the populations of nuclei relax to equilibrium at an exponential rate after the termination of the pulse. Once the magnetization vector is placed onto the y-axis, it rotates in the xy plane at a resonant frequency ultimately decaying back to the z-axis emitting rf radiation over time. This is typically the point of data acquisition (Acq.). A receiver coil resonant at the Larmor frequency, generally located along the x-axis, can detect this rotation called the free induction decay (FID). Fourier transformation of the FID provides the NMR spectrum.
One time constant used to describe this return to equilibrium is called the longitudinal or spin lattice relaxation time (T1). The time (T1) will vary as a function of the magnetic field strength. A second time constant, known as the spin-spin relaxation time (T2), which is due to the exchange of energy between spins, is a description of the return to equilibrium of the transverse magnetization (Mxy) and is always equal to or less than T1.
A spin-echo pulse sequence is normally required to measure T2. The typical pulse-sequence consists of the application of a 90xc2x0 pulse, which results in the rotation of the magnetization to the xy plane, followed by a 180xc2x0 pulse that allows the magnetization to partially rephase producing a non-dephased signal called an echo.
Correlation Spectroscopy (COSY) is a useful technique for determining the signals that may arise from nuclei coupled by a coupling interaction such as a scalar J coupling or a dipole coupling as well as nuclei in close proximity to one another. COSY pulse sequences usually include two 90xc2x0 pulses in succession and gives a signal that varies depending on the time between the application of the two pulses.
Nuclear Overhauser Effect Spectroscopy (NOESY) and Rotational Nuclear Overhauser Effect Spectroscopy (ROESY) have pulse sequences that are used to determine the signal produced from nuclei that are not connected by chemical bonds but are closely oriented in space in the subject molecule.
In general, the practice of high-resolution nuclear magnetic resonance (NMR) spectroscopy yields information about molecular structure and dynamics through the observation of interactions such as chemical shifts and scalar, dipole, quadrupole couplings and the like. These features make in-situ NMR a powerful analytical tool used to study such diverse questions as the metabolism of plants and organisms, the dynamics of geological processes and the characterization of technologically important new materials.
However, there are many circumstances in which it is impractical or undesirable to insert objects or subjects into the bore of a high-field magnet. For many applications it would be useful if a mobile magnet could be scanned over an otherwise inaccessible object or subject in order to acquire magnetic resonance information. The advantage of such ex-situ analysis is that limitations of sample size and transportability no longer prevail. The analysis and imaging of samples that are located outside of the bore of a homogeneous magnet presents serious limitations arising from the presence of strong radio frequency and static field inhomogeneities. As a consequence, the NMR spectra become broadened to the extent that resolution and associated chemical shift information are hidden eliminating the usefulness of ex-situ analysis. The applicability of external detection is limited not only by intrinsic inhomogeneities, but also by the inability to physically rotate many of the samples. Consequently, the use of NMR line narrowing techniques known in the art such as Magic Angle Spinning (MAS), magic angle hopping, dynamic angle spinning or magic angle turning or the like is limited.
Accordingly, a need exists for an ex-situ NMR apparatus and method that can provide chemical shift data from a sample in an inhomogeneous magnetic and rf fields. The present invention satisfies those needs, as well as others, and generally overcomes the deficiencies found in the background art.
The present invention provides an apparatus and methods for NMR spectroscopy and imaging of external samples by refocusing inhomogeneities through the use of correlated, composite z-rotation pulses, producing resolved NMR spectra of liquid and solid samples. By way of example, and not of limitation, in accordance with one aspect of the invention, the observation of chemical shifts is regained through the use of multiple-pulse sequences of trains of composite z-rotation pulses inducing nutation echoes over a region of matched rf and static field gradients.
In one embodiment, the pulse series (xcfx80/2)xe2x88x92y(xcex2({right arrow over (r)}))x(xcfx80/2)y with the constant rotation composite xcfx80/2-pulses tipping the spins back and forth between the xy and yz planes make a position-dependent phase correction possible. In the region of the sample where the static and rf field gradients are matched, a (xcex2({right arrow over (r)}))x pulse applied over the proper time period gives rise to an overall phase shift that exactly reverses the relative positions of (chemically equivalent) fast and slow spins and induces a subsequent nutation echo. Because the same phase shift equally affects nuclei with different chemical shifts, the echo amplitude fully preserves the accumulated phase differences between chemically inequivalent spins. The initial dephasing induced by the pulse (xcex2({right arrow over (r)}))x is refocused during the free evolution (or dwell time) period. A one-point acquisition takes place at the nutation echo maximum, just before the next (xcfx80/2) pulse.
At each step in the pulse train in this embodiment, a phase alternation is performed in order to compensate for undesired evolution during the rf irradiation. The entire rf sequence is repeated, with stroboscopic detection providing the inhomogeneity-free free induction decay. The constant phase shift xcfx8910xcfx84xcex2 remaining at each step of the train can be corrected by sequentially adjusting the phase of the rf pulses or by a shift of the reference frequency in the measured spectrum following data processing.
In another embodiment, the static field (B0) gradient and the rf field (B1) gradient are matched along the sample (x) by a composite xcex2xe2x80x2 pulse sequence that functionally depends from the nutation amplitude xcfx891. By matching the shapes of the rf and static field inhomogeneities, an echo is induced that preserves the chemical shift information. The fact that the obtained resolution is high enough to observe the chemical shift is in most cases synonymous with high resolution NMR and allows the observation of other interactions such as scalar J couplings, dipolar couplings and quadrupolar couplings, which usually require less stringent high-resolution conditions.
In another embodiment, full passage adiabatic pulses are consecutively applied.
In an alternative embodiment, a probe head having magnets creating a characteristic saddle point in the field and with at least one switchable coil is provided. The sample is placed one side of the saddle point with the coils switched off. One preferred pulse sequence begins with an excitation pulse followed by a short period of evolution. The coils are then switched on so that the saddle point is displaced, and the sample lies now on the other side of the saddle point where the field has a complimentary profile (as a simple example: the gradient has an opposite sign). If necessary, the rf frequency can be shifted so that the spins of the sample continue to resonate. A data point acquisition is made after a gradient cycle.
An object of the invention is to provide an apparatus and method for high-resolution ex-situ NMR spectroscopy.
Another object of the invention is to provide a method of observing chemical shift information in an inhomogeneous magnetic field with a pulse sequence of correlated, composite z-rotation pulses that produce resolved NMR spectra of liquid samples.
Another object of the invention is to provide a sequence of radio frequency pulses that have matched radio frequency and static field inhomogeneities to produce an echo that preserves the chemical shift information.
Another object of the invention is to provide a plurality of composite pulses that are configured so that the static field and rf profiles are matched.
Another object of the invention is to provide a NMR probe that has a plurality of permanent magnets to produce a static magnetic field that exhibits a saddle point and a plurality of coils to permit a variable saddle point.
Yet another object of the invention is provide a pulse sequence comprising multiple adiabatic passages.
Another object of the invention is to provide a method for reducing in-situ residual NMR line width due to static field inhomogeneities.
Further objects and advantages of the invention will be brought out in the following portions of the specification, wherein the detailed description is for the purpose of fully disclosing preferred embodiments of the invention without placing limitations thereon.